Genetically modified cells are commonly referred to as transduced cells after having undergone a process commonly referred to as transduction. Transduction can be undertaken with a variety of techniques to allow gene modifying agents to enter the cell. Such genetic modification agents include the use of viral vectors, electroporation, or chemical reagents that increase cell permeability. Transfection and transformation are also common ways to insert genetic material into a cell.
In the case of viral vectors, there are variations on the types used and such types may include lentivirus, retrovirus, adenovirus, or even nanoengineered substances. In the case of electroporation, cells are exposed to a voltage which allows gene modifying agents such as plasmids to enter the cells. A key challenge is to increase the efficiency by which cells are transduced. An efficiency increase can include an improvement in the number of cells transduced within a given cell population or a reduction in the quantity of genetic modification agents needed to genetically alter a given number of cells within a given population.
Lentivirus provides a good example of the advantages and problems associated with cell transduction. Lentivirus is primarily a research tool used to introduce a gene product into in vitro systems. Large-scale collaborative efforts are underway to use lentiviruses to block the expression of a specific gene using RNA interference technology in high-throughput formats. The expression of short-hairpin RNA (shRNA) reduces the expression of a specific gene, thus allowing researchers to examine the necessity and effects of a given gene in a model system. These studies can be a precursor to the development of novel drugs which aim to block a gene-product to treat diseases.
In the field of T cell therapy, an emerging application is to genetically alter T cells in vitro in order to produce chimeric antigen receptors (CARs) which confer the transduced T cells with specificity, typically the specificity of a monoclonal antibody, to a target antigen. In the this manner, a large number of CAR T cells can be generated for use in T cell therapy. The transduction of CAR T cells may also confer cells with enhancement of activation signal, proliferation, production of cytokines and effector function. There is great potential for this approach to improve patient-specific cancer therapy in a profound way. Following the collection of a patient's T cells, the cells are genetically engineered to express CARs specifically directed towards antigens on the patient's tumor cells, then infused back into the patient, where the CAR T cells recognize and kill cancer cells presenting the target antigen.
The object of this invention is to improve the transduction process by increasing the quantity of cells of any given population size that are transduced upon completion of the process, reduce the quantity of gene modification agents used in the process, and/or reduce the cost and complexity of the process, particularly as it relates to transducing T cells.
A common step in the T cell culture and/or T cell transduction process is to use magnetic beads stained with antibody to select a targeted subpopulation of cells from a larger mixed population. For example, a subpopulation of cells such as stem T cells can be selected from a population of leukocytes. Once a subpopulation of cells that recognize the antibody are bound to beads, the entire population is removed from the device and flows past a magnetic field, whereby beads are trapped by the magnetic field, and the subpopulation of cells attached to the beads are thereby isolated from the main population. A significant process simplification would occur if the need to use a flow system to isolate the subpopulation could be eliminated in favor of conducting the process in a static device does not require liquid to flow.